From neural border to migratory stage: A comprehensive single cell roadmap of the timing and regulatory logic driving cranial and vagal neural crest emergence

Neural crest cells exemplify cellular diversification from a multipotent progenitor population. However, the full sequence of molecular choices orchestrating the emergence of neural crest heterogeneity from the embryonic ectoderm remains elusive. Gene-regulatory-networks (GRN) govern early development and cell specification towards definitive neural crest. Here, we combine ultra-dense single cell transcriptomes with machine-learning and large-scale experimental validation to provide a comprehensive GRN underlying neural crest fate diversification from induction to early migration stages in the frog Xenopus tropicalis. During gastrulation, a transient neural border zone state precedes the choice between neural crest and placodes which includes multiple and converging gene programs. Transcription factor connectome and bifurcation analyses demonstrate the early emergence of neural crest fates at the neural plate stage, alongside an unbiased multipotent lineage persisting until after epithelial-mesenchymal transition. We decipher the circuits driving cranial and vagal neural crest formation and provide a broadly applicable strategy for investigating SC transcriptomes in vertebrate GRNs in development, evolution, and disease.